Drug abuse is a severe threat especially to traffic safety, and therefore a number of assays have been developed for convenient testing of drugs of abuse. Many of the tests are immunoassays, and they are mostly competitive immunoassays, which generally are less specific than non-competitive ones. Kerrigan and Phillips have e.g. compared 12 commercially available ELISAs (enzyme linked immunosorbent assays) for opiates, methamphetamine, benzodiazepines, cocaine metabolite, phencyclidine and cannabinoids in whole blood and urine. (Kerrigan S., & Phillips, W., 2001 Clinical Chemistry 47(3):540-547). The test format was competitive immunoassay, and the results were not fully satisfying giving some false results and undesired cross-reactivity.
Drugs of abuse may be assayed not only from blood or urine, but also from saliva, which is the test matrix of choice for roadside testing. Lo Muzio et al. tested a commercial rapid immunological screening test for detecting drugs in urine, and compared two biological matrixes: a nonconventional one, saliva, and a traditional one, urine. They found that saliva specimens were negative in the immunological test for cannabis, THC, benzodiazepines, and tricyclic antidepressants although gas-chromatography-mass-spectrometry (GC-MS) analysis revealed low concentrations thereof, and concluded that the test kit must be improved before being used with saliva (Lo Muzio L., et al., 2005, Int J Immunopathol Pharmacol. 18(3):567-573).
WO 2004/046733 discloses a non-competitive immunoassay for assaying small analytes such as drugs of abuse. The assay uses a first antibody specific for the analyte, and a second antibody specific for an immune complex (IC) between the first antibody and the analyte to improve the sensitivity and specificity of an assay. The anti-IC antibody used was obtained from a naive human antibody fragment phage display library by preincubating the display phages with bound first antibody to sort out those binding to the first antibody as such, whereafter unbound phages were separated and incubated with a mixture of analyte and immobilized first antibody to select phages that bind to the immune complex formed between the immobilized first antibody and analyte, but not to the antibody as such. Very good results for determining morphine in saliva were obtained.
Δ9-tetrahydrocannabinol (Δ9-THC) is the parental drug of cannabis, and it is rapidly metabolized to 11-nor-Δ9-THC-COOH or 11-OH-Δ9-THC that may be determined in urine. Thus the use of cannabis is preferably detected by assaying Δ9-THC in saliva, which is a better indication of recent use than when the metabolites are detected in urine. However, it has turned out to be difficult to develop a quick-test for this analyte, and the test kits on the market are not sensitive enough.
U.S. Pat. No. 6,326,159 and Ullman et al., 1993. Proc. Natl. Acad. Sci 90:1184-1189 describe an immunoassay using anti-IC-antibodies for enhancing the affinity and specificity of primary antibodies. In these papers, a secondary anti-IC antibody, which binds primary anti-analyte antibody that is combined with the analyte, but which does not bind the primary antibody or the analyte alone, has been developed. An antibody that recognizes an immune complex of an antibody to tetrahydrocannabinol (THC) is disclosed. The anti-IC antibody was obtained using an affinity labeled anti-THC antibody as immunogen and selecting an anti-IC antibody the binding of which was enhanced by the presence of Δ9THC. Some cross-reactivity with e.g. THC-metabolites was observed, which is undesirable in forensic drug analysis. The antibodies were prepared by conventional hybridoma technique, which render them too big for application in homogenous immunoassays.
Kintz P. et al. tested a commercial drug device for testing drugs in saliva, and compared the results with GC-MC analysis results, and found that the test device identified only one driver exposed to THC, whereas 18 drivers tested positive with GC-MC. They concluded that a current limitation of the use of this specimen for roadside testing is the absence of a suitable immunoassay that detects the parent compound (THC) in sufficiently low concentrations (Kintz P., et al., 2005. J Anal Toxicol. 29(7):724-727).
The current immunoassays for determining cannabis use are not satisfactory i.a. due to low specificity and sensitivity. Thus there is still a need for an easy, rapid and reliable drug test for cannabis use, which may be used e.g. for roadside testing and preferably for analyzing oral fluid samples, which are non-invasive, easy to perform, and may be achieved under close supervision. The present invention meets at least part of these needs.